Sequence Analysis On The Mitochondrial Genome Of African Rice(Oryza Glaberrima Steud.)

African rice(Oryza glaberrima) is one of the two cultivated rice species. African rice is an important rice germplasm resource. AF-CMS, possessing Oryza glaberrima cytoplasm and African rice introgression lines have been developed in this lab. The mitochondrial genome sequencing has been used effectively in screening genes related to rice cytoplasmic male sterility(CMS). The sequences of twelve rice mitochondrial genomes have been reported,including the Asian rice(Oryza sativa) and the common wild rice(Oryza rufipogon), but African rice mitochondrial genome remains unknown. In this study, we completed sequencing of RAM3 mitochondrial genome, analyzed the sequence by bioinformatics, and developed mitotype-specific sequences markers. The main results are as follows: 1. The sequencing and splicing of RAM3 mitochondrial genome and mitochondrialgenes content. The RAM3 mitochondrial genome was assembled to 519,478 bpsingle circular sequences, with 43.83% G+C content. 91 genes in total were foundamong them, we indentified 66 protein-coding genes, 3 rRNAs and 22 tRNAs,accounting for 9.62% of the mitochondrial genome.In comparison with the reported81 Nipponbare mitochondrial genes, African rice mitochondrial genome has eightspecific ORFs, missing 3 ORFs, a new annotation of 5 ORFs(Nipponbare had butwas not been annotated), 11 genes show difference on the base sequence, a total of 23SNPs, one insertion of three bases. 2. The analysis of the RAM3 mitochondrial genome sequence.(1) We detected 17repeats(>100 bp), then produced three pairs of subgenomic molecules and threeisomers of the MC molecule, respectively produced by recombination of the threedirect repeats and the two inverted repeats.(2) As compared to Nipponbare, IR6888 B,WA-CMS, there are 9 rearrangements in RAM3 mitochondrial genome relative toNipponbare, two specific fragments(NSF1 and NSF2) belong to Nipponbare, twospecific fragments(RSF1 and RSF2) belong to RAM3, seven specific fragments(WSF1 to WSF7) belong to WA-CMS.(3) 6 pairs primer of specific fragments, SG1-SG6, PCR amplification results show that three types(SG1, SG4, SG6)of Sublimonswere detected in Nipponbare, two types(SG2, SG5) of Sublimons were detected bothin RAM3 and Jin23 B, three types(SG2, SG3, SG6) of Sublimons were detected inzhenshan97A, indicated that Sublimons exist in rice mitochondrion. 3. The development of rice MSS markers.45 MSS markers, which were classified in tentypes, is found based on the blastN of four mitochondrial genomes(RAM3,Nipponbare, IR6888 B and WA-CMS), and all these markers were verified in 32different rice materials(18 African rice varieties, 4 rice CMS varieties, 10 NERICAvarieties).Among them, seven, six, six, five MSS markers respectively only belongedto RAM3, Nipponbare, IR6888 B and WA-CMS. These 45 MSS markers can be usedto identify different rice cytoplasmic sources.